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Establishment of Sandwich ELISA for Quality Control in Rotavirus Vaccine Production.

ABSTRACT: Non-replicating rotavirus vaccines are alternative strategies that may improve the protective efficacy of rotavirus vaccines in low- and middle-income countries. The truncated spike protein VP4 (aa26-476, VP4*)was a candidate antigen for the development of recombinant rotavirus vaccines, with higher immunogenicity and protective efficacy compared to VP8* and VP5* alone. This article describes the development of three genotype-specific sandwich ELISAs for P[4], P[6], and P[8]-VP4*, which are important for quality control in rotavirus vaccine production. Our results showed that the detection systems had good specificity for the different genotype VP4* and were not influenced by the E. coli host proteins. Moreover, the detection systems play an important role in determining whether the target protein was contaminated by VP4* proteins of other genotypes. They can also detect the adsorption rate of the adjuvant to the P[4], P[6], P[8]-VP4* protein during the process development. The three detection systems will play an important role in the quality control and process development of VP4* based rotavirus vaccines and facilitate the development of recombinant rotavirus vaccines.

SUBMITTER: Li C

PROVIDER: S-EPMC8876306 | biostudies-literature | 2022 Feb

REPOSITORIES: biostudies-literature

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Establishment of Sandwich ELISA for Quality Control in Rotavirus Vaccine Production.

Li Cao C Luo Guoxing G Zeng Yuanjun Y Song Feibo F Yang Han H Zhang Shiyin S Wang Yingbin Y Li Tingdong T Ge Shengxiang S Xia Ningshao N

Vaccines 20220205 2

Non-replicating rotavirus vaccines are alternative strategies that may improve the protective efficacy of rotavirus vaccines in low- and middle-income countries. The truncated spike protein VP4 (aa26-476, VP4*)was a candidate antigen for the development of recombinant rotavirus vaccines, with higher immunogenicity and protective efficacy compared to VP8* and VP5* alone. This article describes the development of three genotype-specific sandwich ELISAs for P[4], P[6], and P[8]-VP4*, which are impo ...[more]

PMID: 35214701

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Project description:Alpha-Klotho (alphaKl) regulates mineral metabolism such as calcium ion (Ca(2+)) and inorganic phosphate (Pi) in circulation. Defects in mice result in clinical features resembling disorders found in human aging. Although the importance of transmembrane-type alphaKl has been demonstrated, less is known regarding the physiological importance of soluble-type alphaKl (salphaKl) in circulation.The aims of this study were: (1) to establish a sandwich ELISA system enabling detection of circulating serum salphaKl, and (2) to determine reference values for salphaKl serum levels and relationship to indices of renal function, mineral metabolism, age and sex in healthy subjects.We successively developed an ELISA to measure serum salphaKl in healthy volunteers (n=142, males 66) of ages (61.1+/-18.5year). The levels (mean+/-SD) in these healthy control adults were as follows: total calcium (Ca; 9.46+/-0.41mg/dL), Pi (3.63+/-0.51mg/dL), blood urea nitrogen (BUN; 15.7+/-4.3mg/dL), creatinine (Cre; 0.69+/-0.14mg/dL), 1,25 dihydroxyvitamin D (1,25(OH)(2)D; 54.8+/-17.7pg/mL), intact parathyroid hormone (iPTH; 49.2+/-20.6pg/mL), calcitonin (26.0+/-12.3pg/mL) and intact fibroblast growth factor (FGF23; 43.8+/-17.6pg/mL). Serum levels of salphaKl ranged from 239 to 1266pg/mL (mean+/-SD; 562+/-146pg/mL) in normal adults. Although salphaKl levels were not modified by gender or indices of mineral metabolism, salphaKl levels were inversely related to Cre and age. However, salphaKl levels in normal children (n=39, males 23, mean+/-SD; 7.1+/-4.8years) were significantly higher (mean+/-SD; 952+/-282pg/mL) than those in adults (mean+/-SD; 562+/-146, P<0.001). A multivariate linear regression analysis including children and adults in this study demonstrated that salphaKl correlated negatively with age and Ca, and positively with Pi. Finally, we measured a serum salphaKl from a patient with severe tumoral calcinosis derived from a homozygous missense mutation of alpha-klotho gene. In this patient, salphaKl level was notably lower than those of age-matched controls.We established a detection system to measure human serum salphaKl for the first time. Age, Ca and Pi seem to influence serum salphaKl levels in a normal population. This detection system should be an excellent tool for investigating salphaKl functions in mineral metabolism.

Dataset Information

Establishment of Sandwich ELISA for Quality Control in Rotavirus Vaccine Production.

Publications

Establishment of Sandwich ELISA for Quality Control in Rotavirus Vaccine Production.

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